title: ROS-dependent phosphorylation of Bax by wortmannin sensitizes melanoma cells for TRAIL-induced apoptosis.
abstract: The pathways of reactive oxygen species (ROS)-mediated apoptosis induction, of Bax activation and the sensitization of tumor cells for TRAIL (TNF-related apoptosis-inducing ligand)-induced apoptosis are still largely elusive. Here, sensitization of melanoma cells for TRAIL by the PI3-kinase inhibitor wortmannin correlated to the activation of mitochondrial apoptosis pathways. Apoptosis was dependent on Bax and abrogated by Bcl-2 overexpression. The synergistic enhancement was explained by Bax activation through wortmannin, which tightly correlated to the characteristic Bax phosphorylation patterns. Thus, wortmannin resulted in early reduction of the Bax-inactivating phosphorylation at serine-184, whereas the Bax-activating phosphorylation at threonine-167 was enhanced. Proving the responsibility of the pathway, comparable effects were obtained with an Akt inhibitor (MK-2206); while suppressed phosphorylation of serine-184 may be attributed to reduced Akt activity itself, the causes of enhanced threonine-167 phosphorylation were addressed here. Characteristically, production of ROS was seen early in response to wortmannin and MK-2206. Providing the link between ROS and Bax, we show that abrogated ROS production by α-tocopherol or by NADPH oxidase 4 (NOX4) siRNA suppressed apoptosis and Bax activation. This correlated with reduced Bax phosphorylation at threonine-167. The data unraveled a mechanism by which NOX4-dependent ROS production controls apoptosis via Bax phosphorylation. The pathway may be considered for proapoptotic, anticancer strategies.
authors.affiliation: 1] Department of Dermatology and Allergy, Skin Cancer Center, University Medical Center Charité, Berlin, Germany [2] Institute for Chemistry and Biochemistry, Free University of Berlin, Berlin, Germany.
authors.forename: S-A
authors.initials: SA
authors.lastname: Quast
authors.forename: A
authors.initials: A
authors.lastname: Berger
authors.forename: J
authors.initials: J
authors.lastname: Eberle
mesh_terms: D000730:Androstadienes / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D020169:Caspases / Q000378:metabolism; D045744:Cell Line, Tumor; D004789:Enzyme Activation / Q000187:drug effects; D006575:Heterocyclic Compounds, 3-Ring; D006801:Humans; D008545:Melanoma / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology*; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000494:pharmacology*; D000077191:Wortmannin; D051028:bcl-2-Associated X Protein / Q000378:metabolism*
chemical_list: D000730:Androstadienes; D006575:Heterocyclic Compounds, 3-Ring; C548887:MK 2206; D047428:Protein Kinase Inhibitors; D017382:Reactive Oxygen Species; D053221:TNF-Related Apoptosis-Inducing Ligand; D051028:bcl-2-Associated X Protein; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; D051057:Proto-Oncogene Proteins c-akt; D020169:Caspases; D000077191:Wortmannin